Keratoconus (KC) is an abnormal thinning of the human cornea affecting mainly younger individuals. It has an incidence of 1 in 2,000 and is now a leading cause for corneal transplantation in the United States. In the past period the PI established that, in both KC corneas and KC keratocytes, there is an increase in matrix metalloproteinase-2 (MMP-2) activity compared to normal. The hypothesis is that MMP-2 and its associated tissue inhibitors of metalloproteinases (TIMPs) are important in the cascade of events which ultimately results in corneal thinning and that the mechanism(s) which normally regulates MMP-2 activity is altered in the KC cornea. The resulting increase in protease activity affects two areas of the Cornea, the epithelial basement membrane/Bowman's layer and structural components of the extracellular matrix (ECM). An important target of the increased activity may be type VI collagen, which acts as a stabilizing substance between proteoglycans and the collagen fibrils in the stroma. To clarify the role of MMPs in the etiology of KC, the PI proposes to: 1)Determine if there are defects in the structure of MMP-2 and/or TIMPs which would account for the abnormal enzyme activity; 2) Determine if KC corneas have an alteration in TIMP-3 (newly described inhibitor) and/or MT-MMP (cell membrane bound activator of MMP-2); 3) Determine if corneal ECM components, e.g., type XII collagen and proteoglycans, are susceptible to MMP-2; 4) Determine if after cleavage with MMP-2, type VI collagen and other susceptible ECM components interact normally with collagens and proteoglycans; 5) Identify genes which are differentially expressed in KC. The proposed studies will significantly advance our understanding of KC and the changes in gene expression which lead to this disorder. In addition, these studies will also provide basic information on the regulation of MMP activity in the normal cornea and the consequences of protease action on corneal ECM structure and organization.